During adrenocortical cell differentiation changes occur in the smooth endoplasmic reticulum (SER), mitochondria and peroxisomes (Black, '72; Black and Bogart, 73) which can be correlated with the acquisition of steroidogenic capability. The SER proliferates and assumes a complex configuration, mitochondria acquire characteristic shape and cristae morphology, and small peroxisomes connected to the SER increase in number. SER and mitochondria possess most of the enzymes involved in steroid biosynthesis and steroids must shuttle between mitochondria and SER as they traverse the biosynthetic pathway. The organization of enzymes on the membranes of these organelles and the intermembrane relationships which must determine the flow of steroids have not been studied in detail. Peroxisomes are interconnected with the SER but their role in steroid metabolism is unclear. The cells of zona fasiculata and zona reticularis become morphologically distinct during maturation and respond differently to stimulation and suppression (Black, 1975). This may reflect different functional capabilities of the zonal cell types. In this study adrenocortical cells will be examined during differentiation and functional modulation both in vivo and in vitro using combined techniques of electron microscopy, cytochemistry, freeze-fracture, cell fractionation and biochemistry, in order to: 1) Determine if zonal cell types have different functional capabilities; 2) Determine if there are sites of functional specialization in the SER; 3) Establish the route followed by steroids shuttling between SER and mitochondria; 4) Locate enzyme complexes in mitochondrial membranes; 5) Establish the function of peroxisomes and of their relationships with SER.